The present disclosure relates to non-neurotoxin subunits of botulinum neurotoxin complexes, as utilized as a carrier for therapeutic agents. Botulinum toxins are proteins produced by the anaerobic bacterium Clostridium botulinum. There are seven immunologically distinct botulinum neurotoxins, which are designated botulinum neurotoxin serotypes A through G. Each complex consists of a neurotoxin subunit, a non-toxin non-hemagglutinin subunit and hemagglutinin subunits of various sizes and number, depending upon the serotype. The botulinum neurotoxin serotypes vary in the animal species they affect and the severity and duration of paralysis they evoke. The resulting neuroparalytic illness is referred to as botulism. See Preparation and Characterization of Botulinum Toxin Type A For Human Treatment, Schantz, E. J., et al, Therapy with Botulinum Toxin, 1994; v. 25: pp 41-49.
Botulism, or botulinum poisoning, can be caused by exposure to C. botulinum, which can grow in improperly sterilized and sealed foods or from botulinum spores, which are commonly found in soil. Symptoms of botulinum poisoning typically appear 18 to 36 hours after exposure to the bacterium or its spores. Botulinum toxin can pass unattenuated through the lining of the gut and can attack peripheral motor neurons. Symptoms of botulinum poisoning can include difficulty walking, swallowing, speaking, or in more extreme cases, death can result due to paralysis of respiratory muscles. Schantz, E. J. et al., Properties and use of Botulinum Toxin and Other Microbial Neurotoxins in Medicine, Microbiological Reviews, 1992 March, v. 1: pp 80-99.
While passing through the digestive tract, the hydrophobically-bound non-toxin subunits are thought to stabilize and protect the neurotoxin subunit. Protection is especially important for the toxin to pass through the low-pH portions of the digestive tract, which could otherwise denature the neurotoxin subunit. See Preparation and Characterization of Botulinum Toxin Type A For Human Treatment, Schantz, E. J., et al, Therapy with Botulinum Toxin, 1994; v. 25: pp 41-49.
Once the botulinum complex reaches the target neuron, the neurotoxin subunit disassociates from the remaining complex subunits and binds to the presynaptic membrane of the target neuron. The neurotoxin subunit binds to a cell surface receptor and is engulfed via receptor mediated endocytosis. The neurotoxin subunit, which is about 150 kDa, is comprised of a 100 kDa heavy chain portion and a 50 kDa light chain portion, the two portions linked by a disulfide bridge. The 100 kDa heavy chain enables the light chain to bind to the presynaptic membrane of the nerve cell and facilitates the transmembrane transfer of the light chain into the cytoplasm of the cell. The 50 kDa light chain is responsible for the inhibition of acetylcholine release. See Preparation and Characterization of Botulinum Toxin Type A For Human Treatment, Schantz, E. J., et al, Therapy with Botulinum Toxin, 1994; v. 25: pp 41-49.
Once the neurotoxin translocates through an endosomal membrane and enters the cytoplasm of the neuron, the light chain inhibits the release of acetylcholine, which interrupts signals normally transmitted from the nerve cell to neighboring muscle tissue. The result is local paralysis and relaxation of muscle tissue associated with the nerve cell.
Botulinum toxins have been used in the treatment of various neuromuscular disorders and conditions involving muscle spasm, as well as hyperhydrosis, cervical dystonia and blepharospasm.
Botulinum toxin is obtained by growing cultures of C. botulinum in a fermenter under anaerobic conditions, followed by harvesting and purifying the fermented mixture in accordance with known techniques to obtain the botulinum neurotoxin complex.
The protective and stabilizing effects provided by the non-toxin subunits can be useful to facilitate the enteric administration of other therapeutic agents that cannot withstand the extreme conditions of the human digestive tract, or to stabilize a therapeutic agent administered by other routes, such as intramuscular or subcutaneous injection, for example.
The present invention meets this need and provides for a carrier complex for administration of therapeutic agents. In one aspect, an isolated C. botulinum carrier complex, where the carrier complex lacks a native neurotoxin subunit, is provided.